EP0345462B1 - Immunoessai pour des antigènes d'HIV-I utilisant des fragments de F(AB')2 comme sonde - Google Patents

Immunoessai pour des antigènes d'HIV-I utilisant des fragments de F(AB')2 comme sonde Download PDF

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Publication number
EP0345462B1
EP0345462B1 EP89108002A EP89108002A EP0345462B1 EP 0345462 B1 EP0345462 B1 EP 0345462B1 EP 89108002 A EP89108002 A EP 89108002A EP 89108002 A EP89108002 A EP 89108002A EP 0345462 B1 EP0345462 B1 EP 0345462B1
Authority
EP
European Patent Office
Prior art keywords
hiv
probe
fragments
antigens
support
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Expired - Lifetime
Application number
EP89108002A
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German (de)
English (en)
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EP0345462A3 (fr
EP0345462A2 (fr
Inventor
James Lawrence Stewart
Susan K. Ketchum
Robert J. Stumpf
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Abbott Laboratories
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Abbott Laboratories
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Publication of EP0345462A3 publication Critical patent/EP0345462A3/fr
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Publication of EP0345462B1 publication Critical patent/EP0345462B1/fr
Anticipated expiration legal-status Critical
Expired - Lifetime legal-status Critical Current

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
    • C07K16/08—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from viruses
    • C07K16/10—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from viruses from RNA viruses
    • C07K16/1036—Retroviridae, e.g. leukemia viruses
    • C07K16/1045—Lentiviridae, e.g. HIV, FIV, SIV
    • C07K16/1054—Lentiviridae, e.g. HIV, FIV, SIV gag-pol, e.g. p17, p24
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
    • G01N33/56983—Viruses
    • G01N33/56988—HIV or HTLV
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6854—Immunoglobulins
    • G01N33/6857—Antibody fragments
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S436/00—Chemistry: analytical and immunological testing
    • Y10S436/811—Test for named disease, body condition or organ function
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S436/00—Chemistry: analytical and immunological testing
    • Y10S436/811—Test for named disease, body condition or organ function
    • Y10S436/813—Cancer

Definitions

  • the present invention relates to the detection of the Human Immunodeficiency Virus (HIV) in serum, plasma or other body fluids.
  • HIV Human Immunodeficiency Virus
  • this invention describes diagnostic assays which employ F(ab′)2 fragments as a probe for the detection of HIV 1 antigens.
  • HIV 1 is believed to be the causative agent in acquired immunodeficiency syndrome (AIDS) [Chamberland et al., Ann. Int. Med . (1984) 101:617-623; Seligman et al., New Eng. J. Med . (1984) 311:1286-1292].
  • AIDS acquired immunodeficiency syndrome
  • the virus has been isolated from patients with AIDS and AIDS-related complex (ARC) as well as from healthy persons at high risk for AIDS (Gallo et al., Science (1984) 244:500-502).
  • DE-C-3 603 085 discloses the use of tracer-bound antigen to be used for the determination of antigen specific antibodies wherein the tracer is a labeled F(ab′)2 fragment specific to the antigen.
  • Specifically disclosed antigens are viruses of the HTLV III type.
  • HIV 1 Antigen assay for example, the Abbott HIV 1 Antigen assay (Abbott Laboratories, North Chicago, Illinois), have been commercially available on a research basis for detection of HIV 1 antigens. These tests are highly sensitive and provide a direct indication of the presence of the virus. Consequently, detection of HIV 1 antigens may be useful as an aid in the diagnosis and monitoring of HIV 1 infection (Pedersen et al., Brit. Med. J. (1987) 295:567-569; DeWolf et al., Brit. Med. J. (1987) 295:569-572).
  • the sensitivities of the current HIV 1 antigen assays are quite good, however, the specificity varies widely. All manufacturers appear to have samples which nonspecifically react with components of the assay yielding false reactives. For this reason, Abbott provides a confirmatory procedure which involves neutralization of HIV 1 antigen in the sample prior to assaying. Although this increases specificity to near 100%, the cost involved in additional testing warrants efforts to increase the predictive value of antigen testing.
  • the invention described herein provides one method of significantly increasing specificity while also enhancing assay sensitivity and timing.
  • a probe for the detection of HIV 1 antigens in plasma, serum, tissue culture and other biological fluids is employed in the diagnostic assay of the present invention.
  • This probe is a chemically modified antibody. The modification involves enzymic cleavage of antibodies to HIV 1 to produce F(ab′)2 fragments.
  • Immunoassays of the invention comprise:
  • monoclonal antibodies are coated on the solid support to specifically capture HIV 1 p24 antigens.
  • the present invention provides an improved means for the detection of HIV 1 antigens.
  • the use of F(ab′)2 fragments as a probe in an immunoassay to detect HIV 1 antigens increases assay specificity while at the same time enhances assay sensitivity.
  • this probe can be utilized advantageously to improve the immunoassay disclosed in U.S. Patent No. 4,748,110, issued May 31, 1988.
  • any antigen binding fragment such as Fab monomers, produced by cleavage of anti-HIV 1 antibodies with the enzyme papain, can be employed as a probe in the inventive assays.
  • F(ab′)2 fragments specific for the probe are labeled and used to measure the amount of HIV 1 antigen present in the sample.
  • Any label capable of producing a detectable signal can be used in the assays of the invention.
  • Representative labels include enzymes, radioisotopes, fluorescent and chemiluminescent labels.
  • hapten/labeled anti-hapten systems such as a biotin/labeled anti-biotin system can be utilized in the inventive assays.
  • IgG and Igm antibodies may be used in the assays of the invention.
  • Biological samples which are easily tested by the method of the present invention include human and animal body fluids such as whole blood, serum, plasma, cerebrospinal fluid and lymphocyte or cell culture supernatants.
  • Solid supports which can be used in immunoassays of the invention include wells of reaction trays, test tubes, polystyrene beads, strips membranes, microparticles and other solid matrices known to those skilled in the art.
  • reagents for the assays of the invention are ideally suited for preparation of a kit.
  • a kit may comprise carrier means being compartmentalized to receive in close confinement, one or more container means such as vials, bottles, test tubes and the like.
  • Each of the container means comprises one of the separate elements to be used in the assay.
  • Rabbit anti-HIV 1 antibodies were subjected to cleavage with pepsin according to standard procedures (Fanger et al., J. Immunol. (1970) 105:1484-1492; Parham, J. Immunol . (1983) 131:2895-2902).
  • the F(ab′)2 fragments were isolated from the digestion mixture by gel filtration chromatography. Analysis by sodium dodecyl sulfate polyacrylamide gel electrophoresis (Laemmli, Nature (1970) 227:680-685) confirmed the presence of intact F(ab′)2 fragments. This product retains the specificity for recognizing HIV 1 proteins, while eliminating the nonspecific reactions frequently mediated by the Fc portion of the whole molecule antibody.
  • steps 2 and 3 may be combined into one step to facilitate the procedure.
  • Species of antibodies other than human can be used to capture the HIV 1 antigens, and species of F(ab′)2 fragments other than rabbit also can be used as the probe. Better specificity is achieved when the capture and probe antibodies are from different animal species.
  • F(ab′)2 fragments also can be used to coat the beads, in addition to its use as the probe and the conjugate.
  • the incubation times of sample, anti-HIV 1 F(ab′)2 fragments, and conjugate can be reduced to 1 .5 hours, 0.5 hours and 2 hours, respectively, when incubations take place at 40°C in a waterbath.
  • Example 2 The specificity of the assay described in Example 2 was determined using eight nonspecific samples. These samples are repeatably reactive but nonconfirming in the Abbott HIV 1 Antigens assay. This assay uses whole molecule antibodies as a probe. For both assays, an absorbance value which was 0.05 O.D. units greater than that of the negative control was considered the cutoff value. Samples showing higher absorbance values than the cutoff value were considered reactive for HIV 1 antigens. Therefore, samples having a sample to cutoff absorbance value (S/C) greater than or equal to 1.0 were considered reactive. All reactive samples were tested by the Abbott HIV 1 neutralization test, commercially available from Abbott Laboratories for research use. A sample which was neutralized by the procedure was considered to be confirmed positive for HIV 1 antigens.
  • S/C sample to cutoff absorbance value
  • Table 1 shows that all eight specimens were negative using two different production lots of rabbit F(ab′)2 fragments as the probe.
  • S/N signal to noise ratio

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Immunology (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Hematology (AREA)
  • Molecular Biology (AREA)
  • Urology & Nephrology (AREA)
  • Biomedical Technology (AREA)
  • Virology (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • AIDS & HIV (AREA)
  • Organic Chemistry (AREA)
  • Pathology (AREA)
  • General Physics & Mathematics (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Biotechnology (AREA)
  • Cell Biology (AREA)
  • Microbiology (AREA)
  • Analytical Chemistry (AREA)
  • Food Science & Technology (AREA)
  • Biophysics (AREA)
  • Oncology (AREA)
  • Genetics & Genomics (AREA)
  • Tropical Medicine & Parasitology (AREA)
  • Peptides Or Proteins (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Claims (7)

  1. Procédé d'immunoanalyse pour la détection d'antigènes de VIH 1 dans un échantillon biologique, comprenant :
    (a) le revêtement d'un support solide avec un anticorps anti-VIH 1 provenant d'une première espèce animale ;
    (b) la mise en contact du support revêtu avec l'échantillon ;
    (c) la mise en contact du support avec une sonde comprenant des fragments F(ab′)₂ anti-VIH 1 provenant d'une seconde espèce animale ;
    (d) la mise en contact du support avec des fragments F(ab′)₂ marqués spécifiques de la sonde ; et
    (e) la détection du marqueur en tant que mesure de la présence d'un antigène de VIH 1 dans l'échantillon.
  2. Procédé d'immunoanalyse selon la revendication 1, dans lequel ledit marqueur comprend un radioisotope, une enzyme, un composé fluorescent, un composé chimiluminescent ou un élément d'une paire à liaison spécifique.
  3. Procédé d'immunoanalyse selon la revendication 1, dans lequel ledit anticorps anti-VIH 1 appliqué en revêtement sur le support solide comprend au moins un anticorps monoclonal.
  4. Procédé d'immunoanalyse selon la revendication 1, dans lequel ledit anticorps anti-VIH 1 appliqué en revêtement sur le support solide comprend un anticorps polyclonal anti-VIH 1.
  5. Procédé d'immunoanalyse selon la revendication 3 ou 4, dans lequel ledit anticorps anti-VIH 1 appliqué en revêtement sur le support solide comprend en outre des fragments F(ab′)₂.
  6. Procédé d'immunoanalyse selon la revendication 1 pour la détection d'antigènes de VIH 1 dans un échantillon biologique, comprenant :
    (a) le revêtement d'un support solide avec un anticorps anti-VIH 1 provenant d'une première espèce animale ;
    (b) la mise en contact du support revêtu avec l'échantillon, une incubation et un lavage ;
    (c) la mise en contact du support avec une sonde comprenant des fragments F(ab′)₂ anti-VIH 1 provenant d'une seconde espèce animale, une incubation et un lavage ;
    (d) la mise en contact du support avec des fragments F(ab′)₂ marqués spécifiques de la sonde, une incubation et un lavage ;
    (e) la mise en contact du support avec une solution de o-phénylène-diamine-peroxyde d'hydrogène ; et
    (f) la mesure de l'absorbance du produit coloré formé à 492 nm pour déterminer la présence d'antigènes de VIH 1 dans l'échantillon.
EP89108002A 1988-06-10 1989-05-03 Immunoessai pour des antigènes d'HIV-I utilisant des fragments de F(AB')2 comme sonde Expired - Lifetime EP0345462B1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US07/204,799 US4983529A (en) 1988-06-10 1988-06-10 Immunoassay for HIV-I antigens using F(AB')2 fragments as probe
US204799 1988-06-10

Publications (3)

Publication Number Publication Date
EP0345462A2 EP0345462A2 (fr) 1989-12-13
EP0345462A3 EP0345462A3 (fr) 1991-07-31
EP0345462B1 true EP0345462B1 (fr) 1995-04-19

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EP89108002A Expired - Lifetime EP0345462B1 (fr) 1988-06-10 1989-05-03 Immunoessai pour des antigènes d'HIV-I utilisant des fragments de F(AB')2 comme sonde

Country Status (6)

Country Link
US (1) US4983529A (fr)
EP (1) EP0345462B1 (fr)
JP (1) JP2905218B2 (fr)
CA (1) CA1340109C (fr)
DE (1) DE68922243T2 (fr)
ES (1) ES2074062T3 (fr)

Families Citing this family (26)

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US5254457A (en) * 1989-01-11 1993-10-19 The United States Of America As Represented By The Secretary Of The Department Of Health And Human Services Monoclonal antibodies and method for identifying different aids-related viruses
CA2038260A1 (fr) * 1990-05-11 1991-11-12 John C. Mauck Utilisation d'un fragment d'antigene marque par un enzyme dans la determination d'antigenes de chlamydia ou de gonocoques
US6274325B1 (en) * 1990-06-25 2001-08-14 Boehringer Mannheim Gmbh Method for carrying out a homogeneous-immunoassay based on agglutination
CA2086531A1 (fr) * 1990-07-03 1992-01-04 Ebo S. Bos Compose immunoreactif
AT402674B (de) * 1993-01-18 1997-07-25 Waldheim Pharmazeutika Gmbh Verfahren und mittel zum nachweisen von verfahren und mittel zum nachweisen von antigenen antigenen
US6319665B1 (en) 1994-06-07 2001-11-20 Inverness Medical Technology, Inc. Home test kit and method with telephone verification of results
US5874216A (en) 1996-02-23 1999-02-23 Ensys Environmental Products, Inc. Indirect label assay device for detecting small molecules and method of use thereof
EP1255995A2 (fr) 2000-02-16 2002-11-13 Wisconsin Alumni Research Foundation Procede et appareil permettant de detecter des pathogenes microscopiques
EP1423749B1 (fr) 2001-09-04 2009-08-05 Wisconsin Alumni Research Foundation Mecanisme de commutation a cristal liquide
US7807348B2 (en) 2002-03-20 2010-10-05 Wisconsin Alumni Research Foundation Optical imaging of nanostructured substrates
US7125592B2 (en) 2002-04-10 2006-10-24 Wisconsin Alumni Research Foundation Detecting interactions at biomimetic interfaces with liquid crystals
WO2004044583A1 (fr) 2002-11-08 2004-05-27 Wisconsin Alumni Research Foundation Surfaces presentant des gradients de topographie superficielle
US7303694B2 (en) 2003-07-17 2007-12-04 Wisconsin Alumni Research Foundation Liquid crystals with reduced toxicity and applications thereof
JP2007114070A (ja) * 2005-10-20 2007-05-10 Pentax Corp プレート、判定キットおよび判定方法
JP2007114069A (ja) * 2005-10-20 2007-05-10 Pentax Corp 判定方法および判定キット
EP2239575A4 (fr) 2007-12-28 2013-01-02 Sysmex Corp Réactif pour détecter un antigène de vih-1 et procédé de détection
EP2550362B1 (fr) 2010-03-25 2017-01-04 Oregon Health&Science University Glycoprotéines du cmv et vecteurs recombinés
PT2691530T (pt) 2011-06-10 2018-05-10 Univ Oregon Health & Science Glicoproteínas e vectores recombinantes cmv
US20130189754A1 (en) 2011-09-12 2013-07-25 International Aids Vaccine Initiative Immunoselection of recombinant vesicular stomatitis virus expressing hiv-1 proteins by broadly neutralizing antibodies
US9402894B2 (en) 2011-10-27 2016-08-02 International Aids Vaccine Initiative Viral particles derived from an enveloped virus
ES2631608T3 (es) 2012-06-27 2017-09-01 International Aids Vaccine Initiative Variante de la glicoproteína Env del VIH-1
US20150065381A1 (en) 2013-09-05 2015-03-05 International Aids Vaccine Initiative Methods of identifying novel hiv-1 immunogens
EP2873423B1 (fr) 2013-10-07 2017-05-31 International Aids Vaccine Initiative Trimères de glycoprotéines d'enveloppe du vih -1 soluble
EP3069730A3 (fr) 2015-03-20 2017-03-15 International Aids Vaccine Initiative Trimères de glycoprotéines de l'enveloppe du vih-1 soluble
EP3072901A1 (fr) 2015-03-23 2016-09-28 International Aids Vaccine Initiative Trimères de glycoprotéines de l'enveloppe du vih-1 soluble
US9925258B2 (en) 2015-10-02 2018-03-27 International Aids Vaccine Initiative Replication-competent VSV-HIV Env vaccines

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Also Published As

Publication number Publication date
DE68922243D1 (de) 1995-05-24
ES2074062T3 (es) 1995-09-01
JPH0232261A (ja) 1990-02-02
DE68922243T2 (de) 1995-10-26
EP0345462A3 (fr) 1991-07-31
EP0345462A2 (fr) 1989-12-13
JP2905218B2 (ja) 1999-06-14
US4983529A (en) 1991-01-08
CA1340109C (fr) 1998-11-03

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